The most common dominantly inherited ataxia, Spinocerebellar Ataxia Type 3 (SCA3) is also one of nine neurodegenerative diseases caused by polyglutamine expansion. Although polyglutamine diseases share common features centered on protein misfolding, it is increasingly clear that the pathogenesis of SCA3 and other polyglutamine diseases depends greatly on the specific protein context in which the expansion occurs. The studies proposed here extend our longstanding efforts to understand both the disease mechanisms underlying SCA3 and the normal and disease-related functions of the SCA3 disease protein, a de-ubiquitinating enzyme called ataxin-3. The four proposed aims will investigate key unanswered questions in SCA3 and establish important new model systems. Aim 1 will test whether ataxin-3 normally suppresses polyglutamine neurodegeneration by virtue of its activity as a ubiquitin chain editing enzyme and whether polyglutamine expansion in ataxin-3 alters this activity. Aim 2 will address structure-function relationships for this apparently unique enzyme, building on our recent discovery that mono-ubiquitination of ataxin-3 directly enhances its enzymatic activity. Aim 3 will define the basis of early electrophysiologic changes in the cerebellum of SCA3 transgenic mice and test whether these changes can be modified by channel modulators. Finally, Aim 4 will complete our development of SCA3 knock-in mice, bringing to the research field a genetically precise mouse model of SCA3 that is clearly needed. Together these studies are expected to provide new understanding of the biochemical and cellular processes by which SCA3 occurs and identify potential routes to therapy for this fatal neurodegenerative disease.